Cryo-electron microscopy of vitrified biological samples can provide three-dimensional reconstructions of macromolecules and organelles within bacteria and cells at nanometer scale resolution, even in native conditions. Localization of specific structures and imaging of cellular dynamics in cellular cryo-electron microscopy is limited by (i) the use of cryo-fixation to preserve cellular structures, (ii) the restricted availability of electron dense markers to label molecules inside cells and (iii) the inherent low contrast of cryo electron microscopy. These limitations can be mitigated to a large extend by correlative light and electron microscopy, where the sample is imaged by both light and electron microscopy. Here we present a Microscopy and Vitrification Integrated System (MAVIS) that combines a light microscope with a plunger to vitrify thin specimens. MAVIS provides the capability for fluorescence light microscopic imaging of living cells and bacteria that are adhered to an electron microscopy grid and subsequent vitrification within a time frame of seconds. The instrument allows targeting of dynamic biological events in time and space by fluorescence microscopy for subsequent cryo light and electron microscopy. Here we describe the design and performance of the MAVIS, illustrated with biological examples.
Keywords: Correlative microscopy; Cryo-microscopy; Electron microscopy; Light microscopy.
© 2013 Published by Elsevier B.V.